Choleragen activates adenylate cyclase by catalyzing the ADP-ribosylation of guanyl nucleotide-binding regulatory components (G) of adenylate cyclase. The ADP-ribose linkage was stable for 30 degree C for 2 h with or without GTP, whereas GTP was required to stabilize the activity of G. A transferase purified from turkey erythrocytes catalyzed the NAD-dependent ADP-ribosylation of multiple soluble proteins from bovine thymus. Nucleoside triphosphates stimulated this process (ATP more than ITP = GTP more than CTP = UTP). It appears that ADP-ribosylation of cellular proteins by endogenous ADP-ribosyltransferases may be subject to regulation by nucleoside triphosphates. Diethyl pyrocarbonate rapidly inactivated the turkey transferase at 0-4 degree C. Conditions known to activate the enzyme almost doubled the rate constant for inactivation. Enzyme inactivation was dependent on pH. Protection from diethyl pyrocarbonate inactivation could only be achieved in the presence of a complete dinucleotide. Nicotinamide, ADP-ribose, or arginine, or arginine-like compounds did not alter the rate constant for inactivation. A new NAD:arginine ADP-ribosyltransferase was purified from turkey erythrocytes. This enzyme, unlike the transferase previously purified from turkey erythrocytes, was unaffected by chaotropic salts or histone.